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Abstract

Mycobacterium tuberculosis (MTB) is the bacterial pathogen responsible for tuberculosis, a human pulmonary infectious disease. Mycobacterium bovis (BCG) is the causative agent of tuberculosis in cattle, and is often used as the vaccine stain in humans. Specific recipes and methods for culture of MTB and BCG are described in this protocol.

ADNaCl
15 g BSA
6 g Dextrose
2.55 g NaCl
Dissolve in 300 ml ddH2O Note: BSA will take some time to go into solution. Filter to sterilize the supplement and store covered in foil at 4 °C. Do not store for more than one month.

7H9 liquid medium
Dissolve 4.7 g of 7H9 powder in 900 ml dH2O
Add 2 ml glycerol and mix well
Autoclave and let cool completely
Store medium at room temperature in the dark Note: Do not use medium that has been stored for longer than a month.
Right before use, add 2.5 ml 20% Tween 80 (filter sterilized) and 10% ADNaCl supplement. At this point the media can be filter sterilized or used directly. Media to which the supplement and Tween have been added should be stored at 4 °C and used within a few weeks.

This work was funded by 5050 project by Hangzhou Hi-Tech District, Funding for Oversea Returnee by Hangzhou City, ZJ1000 project by Zhejiang Province. This protocol was developed in the Cohen Lab, Department of Genetics, Stanford University, CA, USA [Chen et al. (unpublished)].